circ_0035277 Promotes Gastric Cancer Proliferation and Metastasis via miR-576-3p/LIN28B Axis

Circular RNAs (circRNAs) regulate biological processes of human tumours. Gastric cancer is a prevalent disease that presents tumours with high metastasis. This study aimed to understand regulatory function and mechanism of circ_0035277 in gastric cancer development. circ_0035277 level in tissues and cells was assessed using RT-qPCR assay. Function of circ_0035277 was evaluated via CCK-8, colony formation and Transwell assays. Location of circ_0035277 was verified with FISH assay. Potential mechanism of circ_0035277 was examined using bioinformatics and luciferase reporter assays. BALB/c nude mice were utilised to construct the gastric cancer metastasis model with subcutaneous injection. Haematoxylin–eosin staining was performed to measure visible tumour nodules in lung tissues. Results showed that circ_0035277 is significantly up-regulated in tissues and cell lines of gastric cancer, accelerates the deterioration of gastric cancer, is significantly located in the cytoplasm and serves as a sponge to participate in gastric cancer by targeting miR-576-3p. Furthermore, lin-28 homolog B (LIN28B) was a direct target of miR-576-3p and reversed the role of circ_0035277/miR-576-3p on gastric cancer metastasis. In vivo studies have shown that knockdown of circ_0035277 suppresses gastric cancer metastasis. Overall, circ_0035277 accelerated gastric cancer progression and metastasis by regulating miR-576-3p/LIN28B axis. These findings can provide a potential target for the treatment of gastric cancer.

YBX1 Enhances Metastasis and Stemness by Transcriptionally Regulating MUC1 in Lung Adenocarcinoma

Abnormal expression of the transcription factor Y-box-binding protein-1 (YBX1) is associated with the proliferation, migration, aggressiveness, and stem-like properties of various cancers. These characteristics contribute to the tumorigenesis and metastasis of cancer. We found that the expression levels of Mucin-1 (MUC1) and YBX1 were positively correlated in lung adenocarcinoma cells and lung adenocarcinoma tissue. Our retrospective cohort study of 176 lung adenocarcinoma patients after surgery showed that low expression of both YBX1 and MUC1 was an independent predictor of the prognosis and recurrence of lung adenocarcinoma. In lung adenocarcinoma cells, the silencing/overexpression of YBX1 caused a simultaneous change in MUC1, and MUC1 overexpression partially reversed the decreased tumor cell migration, aggressiveness, and stemness caused by YBX1 silencing. Moreover, chromatin immunoprecipitation (ChIP) and dual-luciferase reporter assays proved that MUC1 was the downstream target of YBX1 and that YBX1 bound to the -1480~-1476 position in the promoter region of MUC1 to regulate its transcription. Furthermore, in mouse xenograft models and a lung cancer metastasis model, MUC1, which is downstream of YBX1, partially reversed the decreased number and size of tumors caused by YBX1 silencing. In conclusion, our findings indicated a novel mechanism by which YBX1 promotes the stemness and metastasis of lung adenocarcinoma by targeting MUC1 and provided a combination approach for diagnosis different from traditional single tumor biomarkers to predict patient prognosis and provide clinical treatment targets.

Ginsenoside CK Inhibits TGF-β-Induced Epithelial-Mesenchymal Transition in A549 Cell via SIRT1

Ginsenoside CK is the main metabolite of protopanaxadiol saponins in intestinal bacteria. Previous studies have shown that ginsenoside CK can affect many aspects of tumor development through a variety of mechanisms. However, few studies have reported the antimetastatic effects of ginsenoside CK in non-small-cell lung cancer (NSCLC). In this study, we explored the effect of ginsenoside CK on epithelial-mesenchymal transition (EMT) induced by TGF-β in A549 cells and the potential molecular mechanisms. Our data showed that ginsenoside CK effectively prevented TGF-β-induced EMT, as indicated by the upregulation of E-cadherin and downregulation of vimentin. Furthermore, ginsenoside CK inhibited the metastatic ability of A549 cells in the tail vein lung metastasis model of nude mice. Additionally, ginsenoside CK decreased the expression of silent information regulator 2 homolog 1 (SIRT1) in the inhibition of EMT induced by TGF-β. Moreover, the antimetastatic effect of ginsenoside CK was reversed by SIRT1 overexpression. Generally, our results indicated the antimetastatic effect and underlying mechanism of ginsenoside CK on TGF-β-induced EMT in A549 cells, suggesting that ginsenoside CK can be used as an effective antineoplastic agent.

Imidazole Analogs of Vascular-Disrupting Combretastatin A-4 with Pleiotropic Efficacy against Resistant Colorectal Cancer Models

Specific targeting of the tumoral vasculature by vascular-disrupting agents (VDA), of which combretastatin A-4 (CA-4) is a main representative, has been considered a new therapeutic strategy against multidrug-resistant tumors. In addition, CA-4 and analogs are tubulin-targeting agents and can exert direct antitumor effects by different mechanisms. Herein, we analyzed a series of synthetic CA-4 analogs featuring N-methylimidazole-bridged Z-alkenes with different halo- or amino-substituted aryl rings in vitro and in vivo, focusing on models of colorectal cancer. Combined in vitro/in vivo structure–activity relationship studies using cell lines and xenograft tumors susceptible to VDA-induced vascular damage demonstrated a clear association of cytotoxic and vascular-disrupting activity with the ability to inhibit tubulin polymerization, which was determined by specific substitution constellations. The most active compounds were tested in an extended panel of colorectal cancer (CRC) cell lines and showed activity in CA-4-resistant and chemotherapy-resistant cell lines. The bromo derivative brimamin was then compared with the known fosbretabulin (CA-4P) by activity tests on DLD-1- (multidrug-resistant) and HT29- (CA-4-resistant) derived xenograft tumors. Treatment did not induce pronounced vascular-disrupting effects in these tumors. Histological analyses revealed distinct tumor substructures and vessel compositions of DLD-1/HT29 tumors, which clearly differed from the tumor models susceptible to VDA treatment. Even so, brimamin effectively retarded the growth of DLD-1 tumors, overcoming their resistance to standard treatment, and it inhibited the outgrowth of disseminated HT29 tumor cells in an experimental metastasis model. In conclusion, combretastatin analogous N-methylimidazoles proved capable of inducing vascular-disrupting effects, comparable to those of CA-4P. In addition, they showed antitumor activities in models of drug-resistant colorectal cancer, independent of vascular-disrupting effects.

CircKIF4A Enhances Osteosarcoma Proliferation And Metastasis By Sponging MiR-515-5p And Upregulating SLC7A11

Background Circular RNAs (circRNAs) are forms of non-coding RNAs that have crucial roles in regulation of various biological processes of several malignant tumors. circKIF4A is closely associated with malignant progression of a variety of cancers. However, the molecular mechanisms as well as roles of circKIF4A in osteosarcoma (OS) have not yet been clearly elucidated. Methods We evaluated the expression of circKIF4A in OS. Colony-formation, cell counting kit-8 (CCK-8), transwell and mice metastasis model assays were done to explore the roles of circKIF4A in vitro and in vivo. TargetScan database, double luciferase, quantitative reverse transcription polymerase chain reaction analysis (RT-qPCR), and RNA immunoprecipitation (RIP) were done to investigate the associated molecular mechanisms. Results In both OS cells and tissues, circKIF4A (hsa_circ_0007255) was found to be upregulated. In vitro and in vivo, circKIF4A knockdown markedly suppressed OS proliferation as well as metastasis. circKIF4A enhanced OS growth as well as metastasis by sponging miR-515-5p and by upregulating SLC7A11. Conclusions We identified the biological significance of the circKIF4A-miR-515-5p-SLC7A11 axis in OS cell proliferation and metastasis, which is important in OS monitoring and treatment. More studies on circKIF4A will inform on the diagnostic markers for early OS screening.

Irinotecan/scFv co-loaded liposomes coaction on tumor cells and CAFs for enhanced colorectal cancer therapy

Background Cancer-associated fibroblasts (CAFs), as an important component of stroma, not only supply the “soils” to promote tumor invasion and metastasis, but also form a physical barrier to hinder the penetration of therapeutic agents. Based on this, the combinational strategy that action on both tumor cells and CAFs simultaneously would be a promising approach for improving the antitumor effect. Results In this study, the novel multifunctional liposomes (IRI-RGD/R9-sLip) were designed, which integrated the advantages including IRI and scFv co-loading, different targets, RGD mediated active targeting, R9 promoting cell efficient permeation and lysosomal escape. As expected, IRI-RGD/R9-sLip showed enhanced cytotoxicity in different cell models, effectively increased the accumulation in tumor sites, as well as exhibited deep permeation ability both in vitro and in vivo. Notably, IRI-RGD/R9-sLip not only exhibited superior in vivo anti-tumor effect in both CAFs-free and CAFs-abundant bearing mice models, but also presented excellent anti-metastasis efficiency in lung metastasis model. Conclusion In a word, the novel combinational strategy by coaction on both “seeds” and “soils” of the tumor provides a new approach for cancer therapy, and the prepared liposomes could efficiently improve the antitumor effect with promising clinical application prospects. Graphical Abstract

Quantitative Detection of Disseminated Melanoma Cells by Trp-1 Transcript Analysis Reveals Stochastic Distribution of Pulmonary Metastases

A better understanding of the process of melanoma metastasis is required to underpin the development of novel therapies that will improve patient outcomes. The use of appropriate animal models is indispensable for investigating the mechanisms of melanoma metastasis. However, reliable and practicable quantification of metastases in experimental mice remains a challenge, particularly if the metastatic burden is low. Here, we describe a qRT-PCR-based protocol that employs the melanocytic marker Trp-1 for the sensitive quantification of melanoma metastases in the murine lung. Using this protocol, we were able to detect the presence of as few as 100 disseminated melanoma cells in lung tissue. This allowed us to quantify metastatic burden in a spontaneous syngeneic B16-F10 metastasis model, even in the absence of visible metastases, as well as in the autochthonous Tg(Grm1)/Cyld−/− melanoma model. Importantly, we also observed an uneven distribution of disseminated melanoma cells amongst the five lobes of the murine lung, which varied considerably from animal to animal. Together, our findings demonstrate that the qRT-PCR-based detection of Trp-1 allows the quantification of low pulmonary metastatic burden in both transplantable and autochthonous murine melanoma models, and show that the analysis of lung metastasis in such models needs to take into account the stochastic distribution of metastatic lesions amongst the lung lobes.

PIWI-interacting RNA 57125 restrains clear cell renal cell carcinoma metastasis by downregulating CCL3 expression

Clear-cell renal cell carcinoma is one of the most common tumors disagnosed, with nearly one third of patients diagnosed with metastatic ccRCC. Although an increasing number of studies has revealed that piwi-interacting RNAs are aberrantly expressed in diverse types of cancers, few of them explored the detailed molecular mechanism of piRNAs in carcinogenesis, particularly in ccRCC. In this study, differentially expressed piRNAs associated with ccRCC were selected by using piRNA-sequencing combined with TCGA data analysis, and piR-57125 was identified. PiR-57125 was found remarkably downregulated in ccRCC samples. Functionally, knockdown of piR-57125 promoted migration and invasion of ccRCC, while overexpression of piR-57125 suppressed ccRCC metastasis. In vivo lung metastasis model also confirmed the same results. CCL3 was identified as the direct target of piR-57125 which could potentially reverse the inhibition effect of piR-57125 in ccRCC metastasis. Further study revealed that piR-57125 modulated ccRCC metastasis through the AKT/ERK pathway. These data indicate that piR-57125 restrains ccRCC metastasis by directly targeting CCL3 and inhibiting the AKT/ERK pathway, and could be a potential therapeutic target for ccRCC.

875 AL009, a fusion protein and multi-Siglec inhibitor, repolarizes suppressive myeloid cells and potentiates anti-cancer effects

BackgroundSialic acid-binding immunoglobulin-type lectins (Siglecs) are a family of cell surface receptors expressed predominantly on myeloid cells that function to promote immune tolerance. Tumors increase the expression of sialic acid glycans and co-opt the immunosuppressive effects of Siglecs, driving tumor resident immune cells toward a cancer permissive phenotype. Due to the overlapping expression profile of Siglec family members on myeloid cells, targeting multiple Siglecs is required for robust efficacy. Here, we present data on AL009, an engineered Siglec-9 extracellular domain-Fc fusion molecule that acts as a sialic acid trap and repolarizes suppressive myeloid cells to activate an anti-cancer immune response.MethodsThe ability of AL009 to competitively block various Siglec-Fc fusion proteins was assessed using cultured human myeloid-derived suppressor cells (MDSCs). MDSC repolarization was analyzed by flow cytometry. MDSCs were co-cultured with activated CD8+ T cells with and without exposure to AL009 and functionally assessed for T-cell activation by flow cytometry and ELISA. In vivo tumor models, including the MC38 and E0771 murine syngeneic subcutaneous models and the B16F10 intravenous lung metastasis model, were used to assess AL009 engineered with a murine Fc (AL009m).ResultsAL009 competitively blocks the ability of at least 5 inhibitory Siglec family members to bind their corresponding sialic acid ligands. When incubated with MDSCs, AL009 promotes CD163 and CD206 downregulation and induces proinflammatory chemokine secretion, consistent with a repolarization effect. Further, AL009 potently relieves MDSC suppression of T cells in a co-culture system. In the MC38 and E0771 murine syngeneic subcutaneous tumor models, AL009m inhibits tumor growth as a monotherapy and in combination with the checkpoint inhibitor anti-PD-L1. In addition, AL009m combines with the tumor antigen targeting antibody TRP-1 in the B16F10 intravenous lung metastasis model to reduce tumor burden.ConclusionsAL009 represents a novel approach to targeting the myeloid cell compartment in oncology by directly repolarizing myeloid cells without cell depletion or limiting the targeting to specific suppressive subpopulations. AL009 has the potential to address tumors that are unresponsive or refractory to standard immunotherapies. These data support further development of AL009 in the clinic with IND enabling studies ongoing.